Isolation and characterization of orbivirus genotypic variants

Area of Science:

• Virology
• Molecular Biology
• Genetics

Background:

• Orbiviruses are segmented RNA viruses with diverse serogroups, including epizootic hemorrhagic disease virus and bluetongue virus.
• Viral genome variability, through RNA deletions or rearrangements like concatemers, can influence viral properties and evolution.

Purpose of the Study:

• To characterize orbivirus variants with RNA deletions and concatemeric RNAs.
• To investigate the molecular mechanisms and functional consequences of these genomic alterations in orbiviruses.

Main Methods:

• Isolation and characterization of orbivirus variants with altered RNA segments.
• In vitro translation assays to determine protein products from wild-type and variant RNA segments.
• RNA hybridization, melting, and reannealing techniques to analyze RNA structure and identify deletion sites.
• T1 RNase mapping and molecular weight estimations to characterize concatemeric RNA structures.

Main Results:

• A variant Ibaraki virus RNA 9 segment showed a deletion of approximately 140 base pairs, leading to the production of a truncated 32 K protein instead of the wild-type 38 K VP6 protein.
• Analysis suggested the deletion occurred towards the 3' end of the positive strand of Ibaraki virus RNA 9.
• RNA genome segments 10 (bluetongue virus type 21) and 9 (Bunyip Creek virus) were observed to form dimeric concatemers in a 5'-3' to 5'-3' orientation.
• In vitro translation of these dimeric concatemers produced proteins identical to those from monomeric RNAs.

Conclusions:

• RNA deletions in orbiviruses can lead to altered protein products, potentially affecting viral function.
• The formation of dimeric concatemeric RNAs in orbiviruses may represent an alternative genomic arrangement with implications for viral evolution.
• These findings contribute to understanding the genetic plasticity and evolutionary pathways of orbiviruses.